DESCRIPTION (Verbatim from applicant's abstract): The major focus of this application is to explore the intracellular Ca2+-dependent mechanisms which control the light-induced membrane conductance changes that underlie the spiking patterns of retinal ganglion cells. Multiple types of voltage-gated conductances are involved in the generation of somatic action potentials and the regulation of the frequency of spike firing in ganglion cells. Although light exerts an important influence on the regulation of the frequency of ganglion cell firing by triggering direct, ionotropic excitatory and inhibitory synaptic inputs, it is often overlooked that both ligand- and voltage-activated conductances are subject to modulation by neuromodulators, which alter the electrical characteristics of channels through various intracellular processes. The experimental hypothesis of this application is that certain Ca2+-dependent intracellular processes, triggered by activation of neurotransmitter receptors, modify the input/output relation of ganglion cells by modulating both intrinsic ion channels and synaptically-gated channels. To test my hypothesis, I will study ganglion cells in turtle retinal slices. Both electrically- and synaptically-evoked responses will be recorded, whereby the contribution of excitatory inputs, as well as different types of voltage-gated K+ currents to the spiking patterns of the ON, OFF and ON-OFF ganglion cells will be evaluated. The interrelationship between light-activated excitatory synaptic inputs, intracellular Ca2+-dependent processes, and ganglion cell spiking responses will be investigated by manipulating intracellular Ca2+, and using selective inhibitors of Ca2+-calmodulin-dependent enzymes. To supplement results obtained by patch clamp experiments, Ca2+-imaging technique will be used to test directly whether glutamate raises intracellular Ca2+ in ganglion cells, to identify the type of glutamate receptor responsible for [Ca2+]i increase, and the pathways of [Ca2+ ]i elevation (e.g., influx through receptor- or voltage-activated channels, or release from internal stores). The effect of intracellular Ca2+ on the functioning of glutamate receptors, and on activation/inactivation properties of voltage-gated currents will be studied under whole-cell and perforated versions of patch clamp.